Etxebarria A, Benito-Vicente A, Stef M, Ostolaza H, Palacios L, Martin C
Unidad de Biofísica (CSIC, UPV/EHU) and Departamento de Bioquímica, Universidad del País Vasco, Apdo. 644, 48080 Bilbao, Spain.
Progenika Biopharma, a Grifols Company, Derio, Spain.
Atherosclerosis. 2015 Feb;238(2):304-12. doi: 10.1016/j.atherosclerosis.2014.12.026. Epub 2014 Dec 20.
The LDL receptor (LDLR) is a Class I transmembrane protein critical for the clearance of cholesterol-containing lipoprotein particles. The N-terminal domain of the LDLR harbours the ligand-binding domain consisting of seven cysteine-rich repeats of approximately 40 amino acids each. Mutations in the LDLR binding domain may result in loss of receptor activity leading to familial hypercholesterolemia (FH). In this study the activity of six mutations located in the cysteine-rich repeats of the LDLR has been investigated.
CHO-ldlA7 transfected cells with six different LDLR mutations have been used to analyse in vitro LDLR expression, lipoprotein binding and uptake. Immunoblotting of cell extracts, flow cytometry and confocal microscopy have been performed to determine the effects of these mutations. In silico analysis was also performed to predict the mutation effect.
From the six mutations, p.Arg257Trp turned out to be a non-pathogenic LDLR variant whereas p.Cys116Arg, p.Asp168Asn, p.Asp172Asn, p.Arg300Gly and p.Asp301Gly were classified as binding-defective LDLR variants whose effect is not as severe as null allele mutations.
低密度脂蛋白受体（LDLR）是一种I类跨膜蛋白，对含胆固醇脂蛋白颗粒的清除至关重要。LDLR的N端结构域包含配体结合结构域，该结构域由七个富含半胱氨酸的重复序列组成，每个重复序列约含40个氨基酸。LDLR结合结构域中的突变可能导致受体活性丧失，从而引发家族性高胆固醇血症（FH）。在本研究中，对位于LDLR富含半胱氨酸重复序列中的六个突变的活性进行了研究。
使用具有六种不同LDLR突变的CHO-ldlA7转染细胞来分析体外LDLR表达、脂蛋白结合和摄取情况。对细胞提取物进行免疫印迹、流式细胞术和共聚焦显微镜检查，以确定这些突变的影响。还进行了计算机分析以预测突变效应。
在这六个突变中，p.Arg257Trp被证明是一种非致病性LDLR变体，而p.Cys116Arg、p.Asp168Asn、p.Asp172Asn、p.Arg300Gly和p.Asp301Gly被归类为结合缺陷型LDLR变体，其影响不如无效等位基因突变严重。
